Flow systems for high-speed analysis and sorting have been used by us to elucidate the biological states of complex eukaryotic cell populations and to analyze and sort large numbers of eukaryotic chromosomes. To detect differences between different classes of cells (e.g., G1, G2, S, transformed, neoplastic) or chromosomes, we stain with organic compounds (e.g., acriflavine, ethidium bromide, chromomycin A3) that exhibit binding specificity and emit fluorescence for subsequent detection in flow and high-speed analysis. The overall object of this project is to develop and improve preparative and cytochemical procedures for flow systems analysis of cells and chromosome in suspension. Fluorescence cytochemistry of three types of fluorophor will be studied. A probe for measuring RNA content per cell using flow systems will be sought. The favored approach is to use a covalent fluorescent conjugate of small polynucleotides for in situ hybridization to cellular RNA. Also fluorogenic substrates of intracellular hydrolytic enzymes will be analyzed and the synthesis will be altered to obtain stains which are particular suited to flow systems applications. Presently we are using 4 methoxy beta napthylamine derivatives as fluorogenic peptidase substrates. DNA specific stains including chromomycin A3, 7-aminoactinomycin D, and di(t-butyl) proflavine will be used to elucidate chromatin content and structure in cells and chromosomes for flow analysis and sorting.